Pre-exposure prophylaxis (PrEP), in which HIV-1 uninfected persons use oral or topical antiretrovirals to protect against sexual HIV-1 acquisition, is one of the most promising new HIV-1 prevention strategies. It has been hypothesized that PrEP, by aborting infections through direct anti-viral activity, may allow HIV-1 specific immune responses to develop in exposed persons. In non-human primate models, T cell responses in blood recognizing SHIV antigens were achieved half of animals receiving PrEP. A chemotherapeutic effect of PrEP could enhance PrEP's efficacy for preventing HIV-1 acquisition, blunt early HIV-1 replication for PrEP breakthrough infections, improve immunogenicity and efficacy of a partially-effective HIV-1 vaccine, and provide anti-HIV-1 protection after PrEP is discontinued. PrEP clinical trials provide a unique opportunity to characterize anti-HIV-1 immune responses in persons on PrEP (compared directly to those on placebo), explore mechanisms for PrEP protection, and anticipate PrEP as chemovaccination paired with candidate HIV-1 vaccines. We are conducting the largest phase III efficacy trial of PrEP - the Partners PrEP Study, a randomized, double-blind, placebo-controlled, clinical trial of oral tenofovir and emtricitabine/tenofovir PrEP among HIV-1 uninfected partners in 4758 African HIV-1 serodiscordant couples. In July 2011, we demonstrated in the Partners PrEP Study that daily oral PrEP using tenofovir and emtricitabine/tenofovir significantly reduced HIV-1 acquisition risk, by 62% (p=0.0003) and 73% (p<0.0001) respectively. We propose to explore anti-HIV-1 responses in HIV-1 exposed but uninfected persons receiving PrEP, evaluate the relationship between anti-HIV-1 immune responses and protection against HIV-1 acquisition and control of viral replication in breakthrough infections observed during prospective follow-up, and, in an exploratory study, collect and test mucosal samples from persons on PrEP to evaluate immune responses at the site of HIV-1 exposure. Our proposed project is innovative in its use of a randomized, placebo-controlled clinical trial population, allowing unbiased, longitudinal comparisons of the effect of PrEP (versus placebo) on induction of HIV-1-specific immune responses, in the study population of HIV-1 serodiscordant couples, allowing precise definition of the degree of HIV-1 exposure, and in its application of state-of-the-art immunologic assays to study the impact of PrEP on HIV-1 protection and control of viral replication. We will 1) determine the frequency, magnitude, and persistence of anti-HIV-1 innate and adaptive immune responses in persons receiving PrEP compared to placebo, 2) assess the effect of anti- HIV-1 responses on protection against HIV-1 acquisition (controlling for PrEP randomization arm) and control of HIV-1 replication (in those with breakthrough infections in spite of PrEP), and 3) measure anti-HIV- 1 innate and adaptive immune responses in mucosal biopsy samples from HIV-1 uninfected persons on PrEP compared to placebo. Our testing will focus on peripheral blood T cell responses, as these have been found most frequently as a potential correlate of HIV-1 resistance in highly-exposed HIV-1 seronegative persons, and we will also test for peripheral innate immune responses (NK cell phenotyping and functional studies, dendritic cell phenotyping). In mucosal samples, we will test for both adaptive and innate immune responses. Our hypothesis is that PrEP allows for development of host immunity that will provide protection against HIV-1 infection and disease. We have assembled a multinational team with the multidisciplinary HIV-1 expertise necessary to execute the proposed studies. Our findings will be directly relevant to the development of a prophylactic HIV-1 vaccine or a combined PrEP-vaccine strategy that might offer greater protection against HIV-1 than either intervention alone.